rabbit polyclonal anti asic2a  (Alomone Labs)

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: Altered hippocampal TFCP2 and ASIC2a expression with glucose hypometabolism in patients with TLE. ( a ) Patient 4’s pre-surgical assessment results: magnetic resonance imaging (left) was negative, electroencephalography (middle) showed spike waves in the temporal lobe, and fluorodeoxyglucose positron emission tomography (right) revealed hypometabolic lesions in the right hippocampus. ( b ) Representative western blot assays of hippocampal TFCP2 and ASIC2a expression in patients with TLE (n = 13) and control patients (n = 10). β-actin was used as a loading control. ( c , d ) Normalised densitometry bar graphs of TFCP2 and ASIC2a for the control subjects and patients with TLE. The experiments were repeated at least 3 times. Data are presented as means ± standard errors and were analysed using unpaired t-tests. *P < 0.05, **P < 0.01 compared to controls. Uncropped western blot images are shown in Supplementary Fig. . Abbreviations, ASIC2a: acid-sensing ion channel 2a; TFCP2: transcription factor CP2; TLE: temporal lobe epilepsy.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Expressing, Magnetic Resonance Imaging, Positron Emission Tomography, Western Blot, Control

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: Altered hippocampal TFCP2 and ASIC2a expression with glucose hypometabolism in pilocarpine-treated rats. ( a ) Representative coronal view microscopic positron emission tomography images in the different phases of epileptogenesis. ( b ) Hippocampal glucose uptake in the different phases after pilocarpine injection. ( c ) Representative western blot assays of hippocampal TFCP2 and ASIC2a expression at different post-seizure time points. ( d ) Bar graph showing decreased hippocampal TFCP2 expression in pilocarpine-treated rats. ( e ) Bar graph showing increased hippocampal ASIC2a expression in pilocarpine-treated rats. The experiments were repeated at least 3 times with at least 3 rats in each group. Data are presented as means ± standard errors and were analysed using 1-way ANOVA and Dunnett’s multiple comparisons test. *P < 0.05 compared with controls, **P < 0.01 compared with controls. Uncropped western blot images are shown in Supplementary Fig. . Abbreviations, ASIC2a: acid-sensing ion channel 2a; TFCP2: transcription factor CP2; SE: status epilepticus; Con: control.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Expressing, Positron Emission Tomography, Injection, Western Blot, Control

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: Glucose deficiency influenced TFCP2 and ASIC2a expression in PC12 cells. ( a ) Representative immunoblot and densitometric analyses showing that cells grown in low-glucose media had significantly decreased TFCP2 expression and significantly increased ASIC2a expression relative to those grown in high-glucose media after 12 and 24 h of growth. ( b ) Representative immunoblot and densitometric analyses showing that cells grown in no-glucose media had significantly decreased TFCP2 expression and significantly increased ASIC2a expression relative to those grown in high-glucose media after 6, 12, and 24 h of growth. ( c ) Representative immunoblot and densitometric analyses showing that STF-31-treated PC12 exhibited significant downregulation of TFCP2 and significant upregulation of ASIC2a relative to DMSO-treated control cells. ( d ) Representative immunoblot and densitometric analyses showing TFCP2 and ASIC2a expression in 2-deoxy-D-glucose-treated PC12 cells cultured in no-glucose media. The experiments were repeated at least 3 times. Data are presented as means ± standard errors and were analysed using 1-way ANOVA and Dunnett’s multiple comparisons test. *P < 0.05, **P < 0.01 compared to controls; # P < 0.05, ## P < 0.01 compared to the DMSO group. Uncropped western blot images are shown in Supplementary Fig. . Abbreviations, ASIC2a: acid-sensing ion channel 2a; TFCP2: transcription factor CP2; DMSO: dimethyl sulfoxide; 2-DG: 2-deoxy-D-glucose; Con: control.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Expressing, Western Blot, Control, Cell Culture

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: TFCP2 inversely regulates ASIC2a expression. ( a ) Representative immunoblot and densitometric analyses showing that TFCP2 siRNA-transfected PC12 cells had 27% lower TFCP2 expression than negative siRNA-treated cells and consequently elevated ASIC2a expression. (b ) Representative immunoblot and densitometric analyses showing that TFCP2 OE plasmid-transfected PC12 cells had 1.76 ± 0.45-fold greater TFCP2 expression than negative control cells and consequently suppressed ASIC2a expression. The experiments were repeated at least 3 times. Data are presented as means ± standard errors and were analysed using unpaired t-tests. *P < 0.05 compared to negative controls. Uncropped western blot images are shown in Supplementary Fig. . Abbreviations, ASIC2a: acid-sensing ion channel 2a; TFCP2: transcription factor CP2; siRNA: short interfering RNA; OE: overexpression; KD: knockdown; NC: negative control.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Expressing, Western Blot, Transfection, Plasmid Preparation, Negative Control, Small Interfering RNA, Over Expression, Knockdown

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: Cellular localisation of TFCP2 and ASIC2a in the epileptic rats’ CA1 regions and glucose-deficient cells. (a ) Double immunofluorescence labelling for TFCP2 and ASIC2a in the epileptic rats’ CA1 regions during acute and latent post-seizure phases. ( b ) Cellular localisation of TFCP2 and ASIC2a expression in PC12 cells grown for 24 h in high-glucose media (control), low-glucose media, no-glucose media, and STF-31 (10 µM)-loaded media. Proteins were probed with anti-ASIC2a (green) and anti-TFCP2 (red) antibodies. Nuclei were counterstained with DAPI (blue). Scale bars = 20 μm. Abbreviations, ASIC2a: acid-sensing ion channel 2a; TFCP2: transcription factor CP2; DAPI: 4′,6-diamidino-2-phenylindole.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Immunofluorescence, Expressing, Control

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: Changes in ASIC2a expression affected the intrinsic excitability of CA1 pyramidal neurons. ( a ) Confocal images of CA1 pyramidal neurons expressing GFP (green) and labelled with neurobiotin (blue). Scale bar = 10 μm. ( b ) Representative traces of action potential firing in response to 200 pA current injections in CA1 pyramidal neurons of the negative control, ASIC2a overexpression, and ASIC2a knockdown groups, respectively. ( c ) Number of action potentials in CA1 pyramidal neurons from the various groups at different current injection steps. Data are presented as means ± standard errors and were analysed using 1- or 2-way ANOVA and Dunnett's multiple comparisons test. *P < 0.05, ASIC2a OE group compared with negative control group; # P < 0.05, ASIC2a KD group compared with negative control group. Abbreviations, ASIC2a: acid-sensing ion channel 2a; GFP: green fluorescent protein; OE: overexpression; KD: knockdown.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Expressing, Negative Control, Over Expression, Knockdown, Injection

Journal: Scientific Reports

Article Title: Glucose Deficiency Elevates Acid-Sensing Ion Channel 2a Expression and Increases Seizure Susceptibility in Temporal Lobe Epilepsy

doi: 10.1038/s41598-017-05038-0

Figure Lengend Snippet: Hippocampal ASIC2a overexpression increased seizure susceptibility. ( a ) Representative images showing GFP immunoreactivity in the hippocampal CA1 region following adeno-associated virus (AAV) vector infusion. Scale bar = 100 μm. ( b ) The time interval from pilocarpine injection to Racine IV seizures was significantly shorter in the ASIC2a overexpression group (26.3 ± 1.3 min) than in the negative control AAV group (31.9 ± 1.9) (n = 30 rats/group). ( c ) The proportion of rats with Racine IV seizures after pilocarpine treatment was significantly higher in the ASIC2a overexpression group (93.3 ± 3.33%) than in the negative control AAV group (66.7 ± 3.33%). Data are presented as means ± standard errors and were analysed using unpaired t-tests or Chi-square tests. *P < 0.05 compared with negative control group. Abbreviations, ASIC2a: acid-sensing ion channel 2a; GFP: green fluorescent protein; DAPI: 4′, 6-diamidino-2-phenylindole; OE: overexpression.

Article Snippet: The sections were then incubated overnight at 4 °C with a mixture of rabbit polyclonal anti-ASIC2a antibody (1:100, Abcam) and mouse monoclonal anti-TFCP2 antibody (1:100, BD Biosciences).

Techniques: Over Expression, Virus, Plasmid Preparation, Injection, Negative Control